Natural products are substances produced by microbes, plants, and other organisms. Microbial natural products offer an abundant source of chemical diversity, and there is a long history of utilizing natural products for pharmaceutical purposes. Despite the emphasis on natural products for human therapeutics, where more than 50% are derived from natural products, only 11% of pesticides are derived from natural sources. Nevertheless, natural product pesticides have a potential to play an important role in controlling pests in both conventional and organic farms. Secondary metabolites produced by microbes (bacteria, actinomycetes and fungi) provide novel chemical compounds which can be used either alone or in combination with known compounds to effectively control insect pests and to reduce the risk for resistance development. There are several well-known examples of microbial natural products that are successful as agricultural insecticides (Thompson et al., 2000; Arena et al., 1995; Krieg et al. 1983).
The development of a microbial pesticide starts with the isolation of a microbe in a pure culture. It then proceeds with efficacy and spectrum screening using in vitro, in vivo or pilot scale trials in a greenhouse and in the field. At the same time, active compounds produced by the microbe are isolated and identified. For the commercialization of a microbial pesticide, the microbe has to be economically produced by fermentation at an industrial scale and formulated with biocompatible and approved additives to increase efficacy and to maximize the ease of application as well as storage stability under field conditions.
As farmers look to expand their insecticide arsenal and as new microbial products are placed on the market, there is a potential for a variety of interactions to occur between new and old insecticides. Combinations of 2 or more insecticides applied to a single crop simultaneously or sequentially have often been used. To address these concerns, scientists have examined the interaction of oils, fungi, and chemical pesticides against pest and beneficial insects using topical and feeding methods (see, for example, Chalvet-Monfray, Sabatier et al. 1996; Meunier, Carubel et al. 1999; Hummelbrunner and Isman 2001; Wirth, Jiannino et al. 2004; Farenhorst, Knols et al. 2010; Shapiro-Ilan, Cottrell et al. 2011); however, not all interactions have yet been studied.
Chromobacterium 
The Beta-Proteobacterium strain, Chromobacterium subtsugae, exhibits insecticidal activity against a wide variety of insects (Martin, Blackburn et al. 2004; Martin 2004; Martin, Gundersen-Rindal et al. 2007; Martin, Hirose et al. 2007; Martin, Shropshire et al. 2007). The mode of action appears to be a combination of antifeedant and toxin activity, with feeding inhibition observed at sublethal doses (Martin, Gundersen-Rindal et al. 2007). In particular, it has been found that Chromobacterium substugae are effective against adult Colorado Potato Beetle (Leptinotarse decemlineata), adult Western Corn Rootworm (Diabrotica virgifera), adult and larval Southern Corn Rootworm (Diabrotica undecimpunctata), larval Small hive beetle (Aethina tumida), larval Diamondback Moths (Plutella xyllostella), adult and larval Sweet Potato Whitefly (Bernisia tabaci) and adult Southern Green Stinkbug (Nezara viridula). Since the finding of C. substugae by Martin and her coworkers, at least three new species of Chromobacteria have been isolated, and characterized; Young et al. (2008) isolated a novel Chromobacterium species, C. aquaticum, from spring water samples in Taiwan, and Kampfer et al. (2009) isolated two species, C. piscinae and C. pseudoviolaceum, from environmental samples collected in Malaysia.
Secondary Metabolites of the Genus Chromobacterium 
Of all known species of Chromobacteria, C. violaceum is studied the most, and published information on secondary metabolites produced by Chromobacteria is based on studies on C. violaceum only. Duran and Menck (2001) have published a comprehensive review of the pharmacological and industrial perspectives of C. violaceum, a Gram-negative saprophyte from soil and water. It is normally considered nonpathogenic to humans, but as an opportunistic pathogen, it has occasionally been the causative agent for septicemia and fatal infections in humans and animals. C. violaceum is known to produce a purple pigment, violacein, which is a bisindole molecule generated by a fusion of two L-tryptophan molecules in the presence of oxygen (Hoshino et al., 1987; Ryan and Drennan; 2009). Violacein biosynthesis is regulated by quorum-sensing, a common mechanism regulating various other secondary metabolism pathways in Gram-negative bacteria (McClean et al., 1997).
Other known metabolites of C. violaceum summarized by Duran and Menck (2001) include hydrogen cyanide, ferrioxamine E, B-lactamic glycopeptides SQ28,504 and SQ28,546, antibiotics such as aerocyanidin, aerocavin, 3,6-dihydroxy-indoxazene, and monobactam SB-26.180, and an antitumoral depsipeptide FR901228. According to the review article by Duran and Menck (2001), C. violaceum also produces unusual sugar compounds such as extracellular polysaccharides and lipopolysaccharides.
Nematodes and Nematocides
Nematodes are non-segmented, bilaterally symmetric, worm-like invertebrates that possess a body cavity and complete digestive system but lack respiratory and circulatory systems. Their body wall is composed of a multilayer cuticle, a hypodermis with four longitudinal cords, and internal musculature (Chitwood, 2003). Their body contents are mostly occupied by digestive and reproductive systems. Most nematodes are free-living but a smaller number of species are ubiquitous parasites of animals or plants.
Root-knot nematodes (Meloidogyne spp.) parasitize a wide range of annual and perennial crops, impacting both quality and quantity of marketable yields. Nematodes in this genus are considered the most economically important plant parasitic nematodes (Whitehead, 1998) Annual crop losses caused by plant-parasitic nematodes have been estimated to exceed US $100 billion (Koenning et al. 1999), with more than half caused by the genus Meloidogyne. The inoculum in this strain comes from eggs that under favorable conditions hatch to release infective second stage larvae (J2s), which migrate in the soil towards a host plant root. Infection occurs through root tip penetration, after which the larvae move to vascular tissue where the nematode becomes sedentary, feeding directly from plant cells. The plant responds by producing giant cells that form galls (root knots). Throughout the reproductive life, females remain imbedded in the plant tissue, and only the egg masses protrude from the root.
The most efficient means for controlling root-knot nematodes is via nematicides that inhibit either egg hatching, juvenile mobility and/or plant infectivity. The development of chemical control for plant-parasitic nematodes is challenging because of both environmental and physiological reasons: 1. Most phytoparasitic nematodes live in a confined area in soil near the roots and hence, delivery of a chemical nematicide is difficult. 2. The outer surface of nematodes is a poor biochemical target, and is impermeable to many organic molecules (Chitwood, 2003). Moreover, delivery of toxic compounds by an oral route is nearly impossible because most plant parasitic nematode species ingest material only after they have penetrated and infected plant roots. Therefore, nematicides have tended to be broad-spectrum toxins with high volatility or with other chemical and physical properties promoting their mobility in soil.
During the past decade, halogenated hydrocarbons (e.g. ethylene dibromide, methyl bromide) have been the most heavily used nematicides around the world. Due to their high human toxicity and detrimental effects on stratospheric ozone layer these compounds were banned in the Montreal Protocol but the use of methyl bromide for nematode and plant pathogen control was extended in the US due to lack of substitution products. Along with organophosphates, carbamates are the most effective non-fumigant nematicides. Unfortunately, most carbamates such as aldicarb and oxamyl are also highly toxic. As of August 2010, the manufacturer of aldicarb, Bayer, has agreed to cancel all product registrations on potatoes and citrus in the US, and aldicarb will be completely phased out by the end of August, 2018. Recently, abamectin—a mixture of two avermectins produced by a soil actinomycete, Streptomyces avermitilis—has been registered for nematicidal use (Faske and Starr, 2006). Syngenta markets this active ingredient as a seed treatment for cotton and vegetables under the trade name Avicta®.
Several microbial plant/nematode pathogens have been reported to be active against plant parasitic nematodes (Guerena, 2006). These biological control agents include the bacteria Bacillus thuringiensis, Burkholderia cepacia, Pasteuria penetrans and P. usgae. Pasteuria Biosciences has launched P. usgae against sting nematodes on turf in the southeastern US. Nematicidal fungi include Trichoderma harzianum, Hirsutella rhossiliensis, H. minnesotensis, Verticillium chlamydosporum, Arthrobotrys dactyloides, and Paecilomyces lilanicus (marketed as BioAct® and Melcon® by Prophyta). Another fungus, Myrothecium verrucaria is available in a commercial formulation, DiTera®, by Valent Biosciences. This is a killed fungus; hence the activity is due to nematicidal compounds. Other commercial bionematicides include Deny® and Blue Circle® (B. cepacia), Activate® (Bacillus chitinosporus) (Quarles, 2005) and an Israeli product BioNem® (Bacillus firmus) (now marketed by Bayer as a seed treatment Votivo®) (Terefe et al. 2009). It has been hypothesized that the detrimental effect of microbial isolates on nematode egg hatching, juvenile mobility and infectivity can be attributed to toxins produced by these organisms (Hallman and Sikora, 1996; Marrone et al, 1998; Siddiqui and Mahmood, 1999; Saxena et al., 2000; Meyer and Roberts, 2002), ability to parasitize or even trap nematodes (Siddiqui and Mahmood, 1996; Kerry, 2001; Jaffee and Muldoon, 1995), induction of systemic resistance (Hasky-Gunther et al. 1998), changing nematode behavior (Sikora and Hoffman-Hergarter, 1993) or interfering with plant recognition (Oostendorp and Sikora, 1990)
Botanical nematicides, such as plant extracts and essential oils, can be used to control nematodes (Kokalis-Burrelle and Rodriguez-Kabana, 2006). Chitwood has summarized the options of using plant-derived compounds for nematode control in his recent review article (Chitwood, 2002). Siddiqui and Alam (2001) demonstrated that potting soil amended with plant parts from the neem tree (Azadirachta indica) and Chinaberry tree (Melia azadirah) inhibited root-knot nematode development of tomatoes. However, no neem products are currently registered in the US for use against nematodes. A new botanical product from Chile (Nema-Q®) based on a Quillaja saponaria tree extract containing saponins (bidesmosidic derivatives of quillajic acid substituted with a trisaccharide at C-3 and an oligosaccharide in C-28) has been recently registered as a an organic nematicide through US EPA and listed for organic farming by the Organic Materials Review Institute (OMRI). It is marketed by Monterey AgResources.
Crop rotation to a non-host crop is often adequate by itself to prevent nematode populations from reaching economically damaging levels (Guerena 2006). Allelochemicals are plant-produced compounds that affect the behavior of organisms in the plant's environment. Examples of nematocidal allelochemicals include polythienyls, glucisonolates, alkaloids, lipids, terpenoids, steroids, triterpenoids and phenolics (Kokalis-Burrelle and Rodriguez-Kabana, 2006; Chitwood, 2002). When grown as cover crops, bioactive compounds from allelopathic plants are exuded during the growing period and/or released to the soil during biomass decomposition. Brassica crops can be used for biofumigation—a pest management strategy based on the release of biocidal volatiles during decomposition of soil-incorporated tissue (Kirkegaard and Sarwar, 1998). However, studies of Roubtsova et al (2007) on the effect of decaying broccoli tissue on M. incognita numbers indicated that for proper control, thorough mixing of plant tissue with the complete nematode-infected soil volume was necessary.
The future of nematode control in agricultural soils relies on two factors: development of nematode resistant crops and the discovery and development of new, broad-spectrum, less toxic nematicides. The cost of research, development and registration of a new chemical nematicides is extremely high (>$200 million), which limits their development. Of the 497 new active ingredients registered for use as a pesticide from 1967 to 1997, only seven were registered as nematicides (Aspelin and Grube, 1999). Besides conventional chemical methods, RNA interference (RNAi) has been proposed as a method for controlling nematodes. Use of gene silencing via RNAi was first demonstrated on Caenorhabditis elegans and quite recently also for plant parasitic nematodes such as Meloidogyne spp. (Bakhetia et al. 2005). The search for new microbial strains to use as sources for biological nematicides is an important goal in order to reduce the significant economic damage caused by plant-parasitic nematodes as well as to reduce the use of toxic compounds currently registered for nematode control.
According to Sasser and Freckman (1987), crop losses by nematodes range from 8 to 20% on major crops around the world. Plant parasitic nematodes can cause considerable crop damage with annual losses estimated at $87 billion worldwide (Dong and Zhang, 2006). Nematode resistant crop varieties and chemical nematicides are currently the main options for nematode control. Fumigants such as methyl bromide are very effective in controlling both soil-borne plant diseases and nematodes but due to the high mammalian toxicity, ozone depleting effects and other residual effects, the use of methyl bromide has already been banned in various countries and its complete withdrawal from the market is planned by international agreement (Oka et al., 2000). Chemical alternatives such as methyl iodide, 1,3-Dichloropropene, and cholorpicrin also have issues with mammalian and environmental safety. Chemical non-fumigant nematicides are being phased out and banned. Most recently, the US-EPA announced that aldicarb was being phased out.